There are significant gaps in our knowledge concerning the relationship of ventilation, gas exchange and body temperature to the mechanisms involved in intracellular acid-base homeostasis. The desirability of developing additional knowledge in this area has been enhanced with the widespread use of hypothermia in human cardiac surgical patients. A new concept of acid- base regulation has been advanced, stemming from work on ectothermic animals, which challenges the traditional view that [H+] is the regulated variable. According to this theory (the alpha imidazole hypothesis), acid- base status is being regulated so that the fractional dissociation of the imidazole moieties of protein histidine remains constant (alphastat regulation). This results in both extracellular and intracellular pH increasing significantly (~0.016U/oC) as a body temperature decreases. Recently, this regimen of acid-base regulation has been advocated for hypothermic cardiac surgical patients in place of the pH-stat regulatory scheme (maintenance of arterial pH at 7.40 during hypothermia). Alphastat regulation is accomplished by varying ventilation and/or inspired gas composition in these patients so that arterial PCO2 decreases with decreasing body temperature. Until now, there has been no way to directly test the validity of this theory since it was impossible to measure the fractional dissociation of imidazole (it had to be calculated). We have developed a method of directly measuring the intracellular fractional dissociation of imidazole in vivo noninvasively and nondestructively using 1H NMR spectroscopy. We propose to use this new method to test the alphastat hypothesis in skeletal muscle cells of ectothermic animals {and in brain and skeletal muscle of mammals}. Additionally, we will test the effects of alphastat and other acid-base regulatory schemes on brain intracellular pH and high energy phosphate levels in mammalian hypothermic animal models using 31P NMR. Comparisons of high energy phosphate levels, brain intracellular pH, {and directly measured alpha-imidazole} to controls for each of the acid-base regulatory regimens should reveal the most appropriate paradigm for hypothermic cardiac surgical patients. The use of these techniques, as indicated by our preliminary results, will reveal much new and exciting information concerning the relationship of hypothermia and ventilatory requirements on an organ system level to intracellular acid- base regulation and cellular high energy phosphate metabolism.